Epigenetic markers are heritable changes in phenotype or gene expression in the absence of changes in DNA sequence. DNA methylation, the most common type of epigenetic modification, plays an important role in cancer, aging, neurodevelopment, and fertility. Recently, changes in global methylation in blood DNA have been associated with environmental exposures, including exposure to persistent organic pollutants, benzene, and air pollution. Little is known about DNA methylation in children, including variations with sex or age and associations with health endpoints or environmental exposures. In this proposal, we will determine whether pre- and/or postnatal exposures to DDT/E and PBDEs, persistent endocrine-disrupting toxicants, are related to global and site-specific DNA methylation in fetal blood and blood of 9-year-old children. We will also determine whether DNA methylation is related to the timing of puberty onset. Ultimately, we aim to identify specific genetic regions where epigenetic modifications can be used as biomarkers of exposure or abnormal pubertal development. We will use the rich collection of biological samples, exposure information, and health outcome data collected by the Center for the Health Assessment of Mothers and Children of Salinas (CHAMACOS) study, a longitudinal birth cohort of 601 women and children with demonstrated high exposure to DDT/E and PBDEs. In this cohort, we will: 1) analyze global DNA methylation in newborn children by pyrosequencing of Alu and LINE-1 repeats, and Infinium Methylation assay;2) determine ontogenetic changes in global methylation in children at birth, 1, 5, 9, and 12 years of age;3) determine the relationship of in utero and 9-year-old blood concentrations of DDT/E and PBDEs with global methylation;4) determine whether global DNA methylation is associated with onset of puberty and hormonal changes;and 5) examine genome-wide site-specific methylation in relation to age, sex, exposure to DDT/E and PBDEs and puberty onset. The total sample size will be 250 children with blood available at birth and 9 years of age. Aims 1, 3, 4 and 5 will be comprised of 100 randomly selected children in a Discovery set and 150 children in a Replication set. Aim 2 will include a subset of 50 children with blood also at 1, 5, and 12 years of age. Epigenetic markers may be a key mechanism by which environmental exposures affect children's health. This study will provide some of the first data on the ontogeny of epigenetic changes in children and their association with environmental exposures and developmental outcomes.